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PDBsum entry 3hcs
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Signaling protein
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PDB id
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3hcs
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References listed in PDB file
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Key reference
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Title
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E2 interaction and dimerization in the crystal structure of traf6.
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Authors
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Q.Yin,
S.C.Lin,
B.Lamothe,
M.Lu,
Y.C.Lo,
G.Hura,
L.Zheng,
R.L.Rich,
A.D.Campos,
D.G.Myszka,
M.J.Lenardo,
B.G.Darnay,
H.Wu.
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Ref.
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Nat Struct Biol, 2009,
16,
658-666.
[DOI no: ]
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PubMed id
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Note: In the PDB file this reference is
annotated as "TO BE PUBLISHED". The citation details given above have
been manually determined.
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Abstract
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Tumor necrosis factor (TNF) receptor-associated factor (TRAF)-6 mediates
Lys63-linked polyubiquitination for NF-kappaB activation via its N-terminal RING
and zinc finger domains. Here we report the crystal structures of TRAF6 and its
complex with the ubiquitin-conjugating enzyme (E2) Ubc13. The RING and zinc
fingers of TRAF6 assume a rigid, elongated structure. Interaction of TRAF6 with
Ubc13 involves direct contacts of the RING and the preceding residues, and the
first zinc finger has a structural role. Unexpectedly, this region of TRAF6 is
dimeric both in the crystal and in solution, different from the trimeric
C-terminal TRAF domain. Structure-based mutagenesis reveals that TRAF6
dimerization is crucial for polyubiquitin synthesis and autoubiquitination.
Fluorescence resonance energy transfer analysis shows that TRAF6 dimerization
induces higher-order oligomerization of full-length TRAF6. The mismatch of
dimeric and trimeric symmetry may provide a mode of infinite oligomerization
that facilitates ligand-dependent signal transduction of many immune receptors.
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Figure 2.
(a) Ribbon diagram of the TRAF6 RZ[1]-Ubc13 complex. The Z[2]
and Z[3] domains are modeled based on superposition of the TRAF6
RZ[1]–Ubc13 complex with the TRAF6 RZ[123] structure and are
shown in gray. (b) Detailed interaction between TRAF6 and Ubc13.
TRAF6 is shown in magenta with the carbon atoms of its side
chains in yellow. Ubc13 is shown in orange with the carbon atoms
of its side chains in gray. (c) Superimposed gel filtration
profiles of Ubc13 mixed with wild-type (WT) or mutant TRAF6
RZ[123] designed to disrupt the interaction. Approximate elution
positions of molecular weight standards are shown. (d) Yeast
two-hybrid experiments on the interaction between Ubc13 and
full-length TRAF2, TRAF3, TRAF5, TRAF6 (positive control) and
its RING mutant C70A (negative control). (e) Superimposed gel
filtration profiles of Ubc13 mixed with wild-type or mutant
TRAF6 RZ[123] with interface residues switched to the
corresponding sequences in other TRAFs: I72A (mutation to the
corresponding TRAF2 sequence), I72K (TRAF3), I72F (TRAF5), L74H
(TRAF3 and TRAF5) and L74R (TRAF2). Approximate elution
positions of molecular weight standards are shown. (f) Promotion
of polyubiquitin chain synthesis by wild-type and mutant TRAF6
RZ[123] in the presence of the E2 complex Ubc13–Uev1A and E1.
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Figure 3.
(a) TRAF6 mutants defective in Ubc13 interaction failed to
rescue IL-1–induced TRAF6 autoubiquitination in
TRAF6-deficient MEFs. The indicated stable cells lines were
treated with IL-1 (1 ng ml^-1) for the indicated times and the
clarified lysates were immunoblotted with the indicated
antibodies. (b) TRAF6 mutants defective in Ubc13 interaction
failed to rescue IL-1–induced IKK activation and I  B
phosphorylation in Traf6^-/- MEFs. IB, immunoblot; WT, wild type.
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The above figures are
reprinted
by permission from Macmillan Publishers Ltd:
Nat Struct Biol
(2009,
16,
658-666)
copyright 2009.
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